Probiotic Tablet Formulations

ABSTRACT

A probiotic tablet comprising a probiotic micro-organism and other nutritionally active ingredients in two zones, a first zone comprising said probiotic micro-organism and precisely selenium, and a second zone comprising at least one said other active ingredient such as iron, other minerals and vitamin B6 kept separated from the probiotic micro-organism of said first zone, the water activity in said probiotic micro-organism containing first zone being no greater than 0.2 and the water content of said tablet being as much as 3% by weight. Good viability of the micro-organisms is obtained despite the relatively high overall moisture content.

The present invention relates to the formulation of probioticmicro-organisms in tablet form. Probiotic micro-organisms areconventionally formulated with other nutritionally active materials suchas vitamins, minerals, carbohydrates, proteins, co-enzymes, enzymes,plant extracts, trace elements, and/or fats. Whilst many probioticmicro-organisms are quite stable when kept by themselves in a driedform, tablet formulations in which the probiotic micro-organisms aremixed with active ingredients of the above kinds are highly unstable.After even brief storage, the recovery of viable micro-organisms uponrehydration of such mixed formulations will be extremely poor.

U.S. Pat. No. 6,254,886 attempts to address this problem by proposingthat the tablet should be in a multilayer form with the probioticmicro-organism being contained in a layer which is free from othernutritionally active materials and which is dry to the extent that itswater content is less than 0.1%. Since water is in fact free to movebetween the different layers of the tablet, this in practice means thatthe carrier material for all the tablet layers has to be dry to thissame extent. Moreover, where large amounts of other active ingredientsare present, they too will have to be aggressively dried if the totalwater content of the probiotic layer is not to rise significantly abovethe limits set in U.S. Pat. No. 6,254,886.

We have now found that the water content in a storage stable probiotictablet formulation can be very much higher than is taught in U.S. Pat.No. 6,254,886 provided that care is taken that the water activity ismaintained below 0.2 (equivalent to 20% relative humidity) and that themixing with the probiotic micro-organisms of certain active materialstaught to be kept separate from the probiotic micro-organisms in U.S.Pat. No. 6,254,886, is not deleterious and may actually improve theviability of the micro-organisms.

The present invention now provides a probiotic tablet comprising aprobiotic micro-organism and other nutritionally active ingredients, thetablet comprising at least two zones, a first of said zones comprisingsaid probiotic micro-organism, and a second of said zones comprising atleast one said other active ingredient kept separated from the probioticmicro-organism of said first zone, the water activity in said probioticmicro-organism containing first zone being no greater than 0.2 and thewater content of said tablet being no less than 0.2% by weight.

Tablets according to the invention, particularly as exemplified belowmay be storage stable at a cool temperature (up to 15° C.) or morepreferably at room temperature (up to 20° C. or more preferably up to25° C.) for several months, e.g. for up to one year or more preferablyup to 18 months or more preferably two years or more. By ‘storagestable’ is meant that after a storage period, the number of viableprobiotic micro-organisms should not have declined by more than a factorof one thousand, preferably not more than one hundred, more preferablynot by a factor of more than 10 e.g., from 5*10⁹ to 5*10⁸, or lesspreferably to 5*10⁷ or still less preferably to 5*10⁶.

According to U.S. Pat. No. 6,254,886, the presence together with theprobiotic micro-organism of other substances valuable in nutritionalphysiology is deleterious. It is suggested that at best there may besome unidentified active materials that are not deleterious. However, wehave found that certain active materials actually improve the stabilityof the product when they are present in the first zone. In accordancewith this, it is preferred that said first zone contains also seleniumas a said at least one other active ingredient. Preferably, said firstzone contains from 1 to 100 μg, e.g. 5 to 75 μg, more preferably 7.5 to60 μg, of selenium, per 10⁹ micro-organisms.

The presence of selenium together with the micro-organisms isparticularly preferred as we have demonstrated that selenium increasesthe storage stability of the tabletted micro-organisms. The mechanismresponsible for this is at present uncertain. It may be that theselenium exerts a beneficial influence in one or more of several waysincluding as a growth medium, as a compression distributor, as astabiliser, as a desiccant or as an antioxidant.

The presence in said first zone of antioxidants generally is alsopreferred. These include ascorbyl palmitate or other ascorbates, propylgallates or other gallates, alpha-tocopherol, magnesium or sodiumsulfite, butylated hydroxyanisole or butylated hydroxytoluene.

Certain active ingredients are however deleterious and should preferablybe excluded from the first zone. These include iron, vitamin B6, vitaminC, zinc, copper, manganese, chromium, pantothenic acid or its salts, andto a lesser extent vitamin B1, so the first zone is preferably free fromamounts of some or all of each of these that axe sufficient materiallyto exert an adverse effect on the stability of the product. Several ofthese materials are available in a micro-encapsulated form. One way inwhich such materials may be present in a tablet according to theinvention without their being present in the first zone is for them tobe encapsulated, but to be present as micro-particles mixed in to theprobiotic micro-organism containing material. If the level of separationimposed by the micro-encapsulation of these materials is not adequate,they may still exert an adverse effect, so we prefer that they shouldnot be mixed into the first zone in micro-encapsulated form, but shouldbe relegated to a more physically distinct and separate macro-region ofthe tablet, such as a distinct layer. This applies especially to ironand copper.

Encapsulated zinc is better tolerated and can be admixed into the firstzone materials.

Vitamin B1 can be present in the first zone in non-encapsulated formwithout much deleterious effect.

Some benefit may come from having certain encapsulated materials mixedinto the first zone. These include micro-encapsulated vitamin B1,micro-encapsulated vitamin B6, micro-encapsulated zinc,micro-encapsulated manganese, micro-encapsulated vitamins A, D, E, B12and B2.

Said second zone preferably contains as at least one said other activeingredient any one of iron, vitamin B6, vitamin C, zinc, copper,manganese, chromium, and pantothenic acid or a salt thereof. Preferablyat least two, more preferably at least four, more preferably at leastsix and preferably all of these are present.

It is preferred that the tablets of the invention have a multi-layerform comprising two or more layers, one of said layers constituting saidfirst zone and another of said layers constituting said second zone.Additional layers may be present. The layers may be formed one over theother or such that a body of material constituting one of the first andsecond zones is enrobed by a layer of material constituting the other ofsaid zones.

Where such a two layer structure is used, it is still possible for thelayer constituting said first zone to contain in encapsulated form somematerials which are required to be kept out of the first zone, but forbetter separation of the probiotic micro-organisms from these materialsit is preferred that they are not present mixed within the first zonelayer but are present only in the second zone. This reduces theinterface area between zones containing the probiotic micro-organism andthese potentially destabilising ingredients. These include particularlyiron, encapsulated iron, vitamin B6, vitamin C, zinc, copper, manganese,chromium, pantothenic acid and its salts, and encapsulated copper and toa lesser degree encapsulated zinc, especially if not stronglyencapsulated, and vitamin B1.

On the other hand, it may be acceptable or even beneficial if mixedwithin the layer constituting the first zone are one, two or anycombination of micro-encapsulated vitamin B1, micro-encapsulated vitaminB6, selenium, micro-encapsulated zinc, iodine, micro-encapsulatedvitamins A, D, E, B12 or B2, nicotinamide, folic acid, or any of theanti-oxidants mentioned herein.

Summing this up, if one were to categorise other active ingredientslikely to be present into three lists: A (aggressive ingredients to bekept well away from the probiotic material, e.g. in a separate layer), B(somewhat aggressive ingredients which are preferably excluded from thefirst zone, but which may well be tolerated either in the first zone orin micro-encapsulated form surrounded by the first zone) and C(non-aggressive or beneficial ingredients that can be present in thefirst zone or if encapsulated can be surrounded by the first zone) theselists would be as follows:

List A

iron

Encapsulated Fe Vitamin B6 Vitamin C Zinc Copper Manganese Chromium

Calcium pantothenateEncapsulated copper

List B Vitamin B1 Nicotinamide List C

Encapsulated vitamin B1Encapsulated vitamin B6

Selenium

Encapsulated zinc

Iodine Magnesium

Encapsulated manganeseEncapsulated vitamin A, D, E, B12, B2Folic acidWhilst not as well tolerated as the above ingredients in List C,nicotinamide may be categorised either in List B or in List C as mayencapsulated zinc.

Whilst layer structures are preferred, it is permissible for the tabletto have a multitude of granules constituting said first zone surroundedby a matrix, wherein said matrix constitutes said second zone or whereinsaid matrix also contains a multitude of granules constituting saidsecond zone.

In order to obtain a low water activity in the first zone, the probioticmicro-organism is preferably mixed with a desiccant carrier materialserving to reduce the water activity of the zone containing theprobiotic micro-organism. Optionally however such a desiccant carriermaterial serving to reduce the water activity of the zone containing theprobiotic micro-organism may be present instead in the second zone.Preferably, such a material is present in both the first and the secondzones. The effect of such a desiccant may be to sequester part of thewater content of the zone so that it is no longer in the form of freewater that can migrate into the probiotic micro-organisms and istherefore prevented from carrying active substances through the cellwalls of such organisms. Such desiccants bind water to specific sites sothat it is no longer able to act as a solvent. These sites include thehydroxyl groups of polysaccharides, the carbonyl and amino groups ofproteins, and others on which water can be held by hydrogen bonding, byion-dipole bonds, or by other strong interactions. Thus, preferreddesiccants include at least one of carboxymethylcellulose, colloidalsilica, polyvinylpyrrolidone, starch, gelatine, hydroxypropylcellulose,microcrystalline cellulose, fumed silicon dioxide, sodiumcroscarmellose, crospovidone, povidone, magnesium aluminium silicate,methylcellulose, sodium alginate, sodium starch glyconate, sodium starchglycolate, gelatine, pregelatinized starch, or sorbitol. The desiccantmay be in particular, a starch selected from corn, rice, or potatostarch, a hydrophilic gum, polysaccharide or galactomannan such aspectin, agar, dextran, maltodextrin, carageenan, tragacanth gum, locustbean gum, acacia gum, guar gum, xanthan gum, ghatti gum, alginic acid orsodium alginate, a cellulose derivative such as methyl cellulose,carboxymethylcellulose, sodium starch glycollate, sodium or calciumcarboxymethylcellulose, hydroxyethyl methylcellulose,hydroxypropyimethylcellulose, ethylhydroxyethylcellulose,ethylmethylcellulose, hydroxyethylcellulose, cellulose acetatephthalate, or microcrystalline cellulose, silica, aluminium silicate,magnesium silicate, aluminium magnesium silicate, sodium silicate orfeldspar, aluminium hydroxide, a protein such as gelatin or casein or apolymer such as acrylate, carboxypolymethylene, a polyalkylene glycol orpolyvinylpyrrolidone. Other steps to reduce the amount of oxygen presentmay be beneficial, including packing under an inert atmosphere such asnitrogen and the use of oxygen barrier packaging materials such asaluminium tubes or high barrier polymers.

The water content of the tablet is at least 0.2% by weight and may beconsiderably higher. Higher water contents remove the need foraggressive drying of materials which may be sensitive to such a process.It is undesirable that the water content in the tablet is too high as itincreases the risk of unforeseen re-crystallisation. Also, it isexpensive to remove water. Thus, the water content can be above 0.5% orabove 1%, but below 6% more preferably below 5%, or 4%, 3%, or even 2%.Alternatively, the water content can be above 0.5% or above 1% or 2%,but below 6% more preferably below 5%, or 4%, or 3%. Alternatively, thewater content can be above 0.5% or above 1% or 2% or 3%, but below 6%more preferably below 5%, or 4%. The water content can go up to 7% byweight.

At the same time, the water activity is preferably below 0.18, morepreferably below 0.15, still more preferably below 0.13, e.g. 0.10, oreven 0.08. The water activity may be still lower, e.g. 0.05 or even0.02. The water activity may lie between 0.2 and any of the foregoingfigures or between any two of them.

Each of the foregoing figures for water activity relate to the firstzone of the tablet. Normally, following internal equilibration, thiswill also be the water activity of the tablet as a whole. Unless aninternal water excluding barrier layer is present separating of thefirst zone, the water activity will equilibrate throughout the tablet toreach the same value throughout.

To improve the separation of the probiotic micro-organisms from theingredients that are hostile to their stability, said first zone may beseparated from said second zone by a water excluding barrier material.Additionally or instead, the tablet as a whole may be surrounded by awater excluding material. Such materials may be cellulose acetatephthalate, methacrylic acid copolymers, alginic acid, zein, modifiedstarch, polyvinylacetate phthallate, hydroxypropylmethylcellulosephthalate, cellulose acetate phthalate, or shellac.

The barrier materials may more preferably be or include a fat basedmaterial, which may be applied by a process of hot melt coating. Theseinclude but are not limited to fatty acid triglycerides, e.g.hydrogenated palm oil or beef tallow and mixtures of triglyceride estersof higher saturated fatty acids along with varying proportions of mono-and di-glycerides, e.g. hard fats.

Tablets according to the invention may be stored in a containercontaining a desiccant for absorbing water so as to reduce the wateractivity in the area surrounding said tablet. Thus, the tablets may bepackaged in such a way as to preserve their initial state of drynesswithin acceptable limits. This may involve packaging the tablets in amoisture impermeable container such as a tube or a blister pack, whichmay contain a desiccant agent such as silica gel. For protection againstoxygen such a pack may contain an oxygen scavenger material such asAmosorb™, ascorbyl palmitate or other ascorbates, propyl galates orother gallates, alpha-tocopherol, magnesium or sodium sulfite, butylatedhydroxyanisole or butylated hydroxytoluene. Oxygen absorbents asdescribed in U.S. Pat. Nos. 5,885,481, 5,744,056, or 6083585 can beused.

The tablets may contain additional materials, especially in the secondzone, such as plant materials, including herb materials, for exampleEchinacea, elderberry extract, blueberry extract, cranberry extract androse hip.

The term ‘probiotic micro-organism’ is well understood by those skilledin the art to which this invention pertains. Probiotics are microorganisms, which in tablet formulations are normally freeze dried andare normally live, which have a beneficial) effect on health wheningested. The probiotic micro-organisms may be lactic acid producingbacteria, e.g. Lactobacilli and Bifidobacteria bacteria. Probioticmicro-organisms that may be present include but are not limited to:

Bibidobacterium

bifidum

longum

adolescentis

animalis

infantis

breve

lactis

Lactobacillus

casei

acidophilus

paracacei

plantarum

rhamnosus

reuteri

gasseri

jensenii

delbruekii including subspecies delbrueckii and bulgaricus

helveticus

salivarius

brevis

johnsonii

crispatus

Bacillus

coagulans

Saccharomyces

boulaardii

cerevisiae

Streptococcus

thermophilus

Enterococcus

faecium

faecalis

Propionebacterium

freudenreichii

Lactococcus

lactis

Propionebacterium

freudenreicii

Each tablet suitably will contain from 10⁶, more preferably from 10⁷ to10¹², e.g. from 10⁸ to 10¹⁰, viable micro-organism cells.

Preferred methods for producing tablets from the tablet ingredientsinclude standard tabletting methods, including those conventionally usedfor producing multi-layer tablets. As we have found that excessivetabletting pressure can decrease the viability of the micro-organisms,we prefer that the compression pressure for the probiotic layer shouldnot exceed 50 kN/cm², corresponding to a tensile strength below 100N(Erweka equipment).

The tablets may be designed to be chewed or to be swallowed whole. Whenthe tablets disintegrate on consumption, whether in the mouth or in thestomach, the micro-organisms are exposed to the materials from whichthey were held separate in the tablet structure. This may harm themicro-organisms if the local concentration of the damaging materials istoo high. To guard against this, it is preferred that the disintegrationof the two zones or layers be spaced in time to a degree to allow thecontents of one zone to be diluted and dispersed before the other zoneis released. This may be achieved by the inclusion in one zone or layerof disintegrant agents selected to provide faster disintegration of thatzone. The effect may be quantitated by a dissolution test in which atablet is allowed to disintegrate in unstirred water in a beaker at 25°C. and after one zone has disintegrated, the remainder of the tablet isremoved, dried and weighed to establish the amount of that zone of thetablet remaining (as a proportion of the total amount of that zoneinitially). Preferably, in such a test, the remainder should amount tono less than 20%, more preferably no less than 50%, most preferably noless than 70% of the original amount of that zone or layer.

The test may alternatively be conducted on a time measurement basis inwhich the tablet is allowed to dissolve as before but the time when afirst zone has disintegrated is noted and the time when the total tablethas disintegrated is noted. If both layers disintegrated at the samerate, the first time period would be the same as the totaldisintegration time. When one zone disintegrates faster, as preferred,the first time period as a percentage of the total disintegration timeis preferably no more than 50%, more preferably no more than 20% andmost preferably no more than 5% of the total.

Ingredients that promote rapid disintegration (super-disintegrants) thatcan be included in one of the zones for this purpose include sodiumcroscarmellose, cross linked sodium carboxymethylcellulose,crospovidone, sodium starch gycolate, sodium starch glyconate andpregelatinized starch.

The invention will be further described with reference to the followingillustrative examples of multilayer tablets, containing freeze driedprobiotic cultures and vitamins/minerals, herbals or drugs.

EXAMPLE 1

The following ingredients were formulated into a two layer tastychewable tablet incorporating lactic acid bacteria, vitamins andminerals using Xylitol and Isomalt to provide bulk and sweetening:

Per tablet:

Vitamin A mcg 700.00 Retinolacetate Vitamin D mcg 5.00 CholecalciferolVitamin E IU 10.43 D,L-alfa-tocopherolacetate Vitamin B1 (salt) mg 1.00Thiaminenitrate Vitamin B2 mg 1.20 Riboflavin Vitamin B6 (salt) mg 1.10Pyridoxine chloride Vitamin B12 mcg 1.40 Cyanocobalamin Nicotinamide mg13.00 Nicotinamide Pantothenic acid mg 5.00 D-Calcium pantothenate Folicacid mcg 100.00 Folic acid Vitamin C mg 60.00 Ascorbic acid Calcium mg200.00 Calcium carbonate Magnesium mg 50.00 Magnesium oxide Iron mg10.00 Ferrous fumarate Zinc mg 7.00 Zinc oxide Copper mg 0.70 Cupricoxide Manganese mg 2.00 Manganese sulfate Chromium mcg 50.00 Chromium(III) chloride Selenium mcg 30.00 Sodium selenate Iodine mcg 90.00Potassium iodide Biotin mcg 30.00 d-Biotin Vitamin K mcg 30.00Phytomenadione Lactobacillus GG cfa 1 × 10⁹The vitamins and minerals (except for selenium) are mixed with thefollowing excipients:

Xylitol 320 mg Microcrystalline cellulose 64 mg Flavour 33 mg Stearicacid 22 mg Silicon dioxide 7 mg Acesulfam potassium 2 mg (in total 700mg)The freeze dried probiotic culture (10 mg=3×10⁹) and the selenium ismixed with:

Isomalt 253 mg Xylitol 100 mg Microcrystalline cellulose 31 mg Magnesiumstearate 4 mg silicon dioxide 2 mg (in total 400 mg)Tablets were produced having two superposed layers using a conventionaltabletting machine, the ingredients of one layer being filled over theingredients of the other.

Tablet weight 1100 mg Tablet size 11 by 16.5 mm oval Water activity** inculture granulate <0.1 Water content* in culture granulate   2% Wateractivity** in tablet  0.09 Water content* in tablet 2.7% **Nova Sina . .. , *Karl FisherFor comparison, a single layer tablet was produced containing the sameingredients. The viability of the micro-organisms was measured afterstorage of the tablets over nine months with the following results:

Months Single layer tablet Dual layer tablet 0 7.3 * 10⁸ 1.5 * 10⁹ 1.56.9 * 10⁷ 1.1 * 10⁹ 6 1.5 * 10⁷ 3.4 * 10⁷ 9  <2 * 10³ 1.1 * 10⁵It can be seen that the two layer tablet of the invention maintained theviability of the micro-organisms over the total storage period better bya factor of over 100.

EXAMPLE 2

The following ingredients were formulated as a two layer tablet toswallow with lactic acid bacteria, vitamins and minerals.Per tablet:

Vitamin D mcg 5.00 Cholecalciferol Vitamin E IU 14.90D,L-alfatocopherolacetate Vitamin B1 (salt) mg 5.00 ThiaminenitrateVitamin B2 mg 5.00 Riboflavin Vitamin B6 (salt) mg 5.00Pyridoxinchloride Vitamin B12 mcg 3.00 Cyanocobalamin Biotin mcg 30.00d-Biotin Nicotinamide mg 18.00 Nicotinamide Pantothenic acid mg 5.00D-Calciumpantothenate Folic acid mcg 400.00 Folic acid Vitamin C mg90.00 Ascorbic acid Magnesium mg 90.00 Magnesium oxide Zinc mg 15.00Zinc oxide Manganese mg 2.50 Manganese sulfate Chromium mcg 30.00Chromium (III) chloride Selenium mcg 50.00 Sodium selenate Iodine mcg100.00 Calcium iodide Lactobacillus GG cfu 1 × 10⁹The vitamins and minerals (except for selenium) are mixed with thefollowing excipients:

Microcrystalline cellulose 58 mg  Magnesium stearate 4 mg Stearic acid 3mg Silicon dioxide 1 mg (in total 555 mg)  The freeze dried probiotic culture (10 mg=3×10⁹) and the selenium aremixed with:

Microcrystalline cellulose 183 mg Magnesium stearate  2 mg Silicondioxide  0.4 mg (in total  195 mg)Tabletting was conducted as in Example 1 and the 2-layer tablets werefilled into aluminium tubes with desiccant in the lid.

Tablet weight 750 mg Tablet size 12 by 4 mm circular Water activity** inculture granulate 0.07 Water content* in culture granulate   2% Wateractivity** in tablet 0.07 Water content* in tablet 3.2% **Nova Sina . .. , *Karl Fisher

EXAMPLE 3

The following ingredients were formulated into a two layer tastychewable tablet incorporating lactic acid bacteria, vitamins andminerals using Xylitol and Lactitol to provide bulk and sweetening:Per tablet:

Vitamin A mcg 700.00 Retinolacetate Vitamin D mcg 5.00 CholecalciferolVitamin E IU 10.43 D,L-alfa-tocopherol acetate Vitamin B1 (salt) mg 1.00Thiaminenitrate Vitamin B2 mg 1.20 Riboflavin Vitamin B6 (salt) mg 1.10Pyridoxine chloride Vitamin B12 mcg 1.40 Cyanocobalamin Nicotinamide mg13.00 Nicotinamide Pantothenic acid mg 5.00 D-Calcium pantothenate Folicacid mcg 100.00 Folic acid Vitamin C mg 60.00 Ascorbic acid Calcium mg200.00 Calcium carbonate Magnesium mg 50.00 Magnesium oxide Iron mg10.00 Ferrous fumarate Zinc mg 7.00 Zinc oxide Copper mg 0.70 Cupricoxide Manganese mg 2.00 Manganese sulfate Chromium mcg 50.00 Chromium(III) chloride Selenium mcg 30.00 Sodium selenate Iodine mcg 90.00Potassium iodide Biotin mcg 30.00 d-Biotin Vitamin K mcg 30.00Phytomenadione Lactobacillus GG cfu 1 × 10⁹The vitamins and minerals (except for selenium) are mixed with thefollowing excipients:

Lactitol 209 mg  Microcrystalline cellulose 39 mg Flavour 2.5 mg Stearic acid 44 mg Silicon dioxide 14 mg Neohesperidin 10% 0.2 mg Citric acid monohydrate  2 mg (in total 1160 mg)  The freeze dried probiotic culture (10 mg=3×10⁹) and the selenium ismixed with:

Lactitol 394 mg  Microcrystalline cellulose 21 mg Stearic acid 14 mg (intotal 440 mg) Tabletting was conducted as in Example 1 and the 2-layer tablets werefilled into aluminium tubes with desiccant in the lid.

Tablet weight 1600 mg Tablet size 16 mm circular Water activity** inculture granulate <0.1 Water content* in culture granulate 3.1% Wateractivity** in tablet 0.09 Water content* in tablet 3.7% **Nova Sina . .. , *Karl FisherThe tablets were tested for stability by storage for 18 months in higher(24%), middle (20%) and lower (7%) relative humidity conditions andviability of the micro-organisms was monitored, with the followingresults:

Months 24% humidity 20% humidity 7% humidity 0 1.7 * 10⁹ 1.7 * 10⁹ 1.7 *10⁹ 6 8.7 * 10⁶ 1.3 * 10⁸ 0.9 * 10⁹ 9 8.5 * 10⁵ 8.0 * 10⁸ 0.6 * 10⁹ 121.4 * 10⁴   3 * 10⁶ 0.6 * 10⁹ 18  <2 * 10³ 5.7 * 10⁵ NDThus, it can be seen that the tablets of the invention providedexcellent long term stability.

In the above Examples, the vitamins used were in some cases supplied inan encapsulated form, others were used in non-encapsulated form. Thetable below indicates the ingredients present in the vitaminformulations used

Active ingredients Amount Vitamin D (Cholecalciferol) 5 mcg = 200 IU AsCholecalciferol Concentrate 2.00 mg Powder (analysed to 110 IU/mg)Cholecalciferol 6 mcg Sucrose 0.68 mg Gelatin 0.42 mg Modified Starch0.42 mg Triglycerides, medium-chain 0.38 mg Butyl Hydroxytoluene 19 mcgSodium Aluminosilicate 3 mcg Water 72 mcg Vitamin E (D-α-tocopherol)14.90 IU As α-Tocopherol Acetate 30.08 mg Concentrate (Powderform)(analysed to 52.5 w/w %) DL-α-Tocopherol Acetate 15.79 mg MaizeStarch 6.02 mg Gelatin 5.11 mg Sucrose 1.41 mg Sodium Aluminosilicate0.39 mg Water 1.35 mg Vitamin B1 (Thiamin) 5 mg As Thiamin Nitrate 33%14.85 mg Thiamin nitrate 4.95 mg Mixture of mono-, di and 9.90 mgtriglycerides Vitamin B2 (Riboflavin) 5 mg As Riboflavine 33% 15.60 mgRiboflavine 5.20 mg Mixture of mono-, di and 8.84 mg triglycerides MaizeStarch 1.56 mg Vitamin B6 (Pyridoxine) 5 mg As Pyridoxine Hydrochloride33% 15.45 mg Pyridoxine Hydrochloride 5.15 mg Mixture of mono-, di and10.30 mg triglycerides Vitamin B12 3 mcg As Cyanocobalamine 0.1% 1.87 mg(analysed to 0.11%) Cyanocobalamine 3 mcg Maltodextrin 2.64 mg Sodiumcitrate 27 mcg Citric acid 20 mcg Water 120 mcg Biotin 30 mcg AsD-Biotin 32 mcg Nicotinamide 18 mg As Nicotinamide 33% 56.16 mgNicotinamide 18.72 mg Mixture of mono-, di and 31.82 mg triglyceridesSilicon dioxide 5.62 mg Pantothenic Acid 5 mg As Calcium Pantothenate5.56 mg Folic Acid 400 mcg Folic Acid 0.49 mg Folic Acid 0.44 mgAbsorbed Water 49 mcg Vitamin C (Ascorbic Acid) 90 mg As Ascorbic Acid97% 100.21 mg Ascorbic Acid 97.20 mg Maize Starch 3.01 mg Vitamin A(Retinol) 700 mcg Vitamin A Concentrate Synthetic 5.21 mg (Powderform)(analysed to 565 IU/mg) Retinol Acetate 1.02 mg Sucrose 1.77 mgGelatin 1.25 mg Modified Starch 0.83 mg Butylated Hydroxytoluene 0.07 mgSodium Aluminosilicate 18 mcg Water 0.25 mg

EXAMPLE 4

Effect of selenium on viability on storage:The following mixtures have been stored in a dehumidified room at atemperature of 25° C. Starting counts and counts of viable organismsafter the indicated storage period were measured.(a)5 mg LGG+295 mg Microcrystalline cellulose:Start week 0: count 3.5×10⁹ Cfu/tabletEnd week 8: count 2.9×10⁹ Cfu/tablet(b)5 mg LGG+0.05 mg Selenium+295 mg Microcrystalline cellulose:Start week 0: count 4.0×10⁹ Cfu/tabletEnd week 8: 4.6×10⁹ Cfu/tabletIt can be seen that the presence of selenium was beneficial to thestability of the micro-organisms, and indeed that the numbers ofrecoverable micro-organisms even increased on storage in the presence ofselenium.In each case the probiotic bacteria were Lactobacillus rhamnosus GG“Grade P” (ATCC 53103) as a concentrated, freeze-dried bacterial powder.

EXAMPLE 5—TABLETS WITH DIFFERENTIAL SPEED OF DISINTEGRATION OF LAYERS

The composition of the probiotic layer, but not of the vitamin/minerallayer, of the tablet of Example 2 was modified in three ways as follows:Freeze dried probiotic culture and selenium—unchanged

Probiotic Layer Formulation (a)

Selenium granulate 2% 2.5 mg Silicon dioxide 0.4 mg Lactose anhydrous181 mg  Magnesium stearate 1.5 mg

Probiotic Layer Formulation (b)

Selenium granulate 2% 2.5 mg Silicon dioxide 0.4 mg Lactose anhydrous171.7 mg  Croscarmellose sodium 1.5 mg

Probiotic Layer Formulation (c)

Selenium granulate 2% 2.5 mg Silicon dioxide 0.4 mg Lactose anhydrous171.7 mg  Magnesium stearate 1.5 mg Povidone 9.3 mgThe dissolution time of the two layers was measured in each case byobserving disintegration of the tablet in a beaker of water with thefollowing results:Vitamin/mineral layer: 14 minutesProbiotic layer:

Formulation (a)  6 minutes Formulation (b) 1 minute 45 sec Formulation(c) 15 sec

EXAMPLE 6—FURTHER TABLETS WITH DIFFERENTIAL SPEED OF DISINTEGRATION OFLAYERS

A two layer tablet was produced in which a probiotic containing layerwas formulated as follows:The freeze dried probiotic culture (10 mg=3×109) is mixed with:

Selenium Granulate 2% 2.5 mg Silicon Dioxide 0.8 mg Magnesium Stearate1.5 mg Cellulose, Microcrystalline Cellulose 152.4 mg  Hypromellose15000 27.8 mg The vitamin/mineral layer was as from example 2 with either 0%Croscarmellose (Formulation 1)

or 5% Croscarmellose Sodium (Formulation 2)

In a dissolution test conducted as above, the results were as follows:Disintegration timeProbiotic layer 10 minutesVitamin/mineral layer1: 37 minutes2: 3 minutes

1. A probiotic tablet comprising a probiotic micro-organism and othernutritionally active ingredients, the tablet comprising at least twozones, a first of said zones comprising said probiotic micro-organism,and a second of said zones comprising at least one said other activeingredient kept separated from the probiotic micro-organism of saidfirst zone, the water activity in said probiotic micro-organismcontaining first zone being no greater than 0.2 and the water content ofsaid tablet being no less than 0.2% by weight.
 2. A tablet as claimed inclaim 1, wherein said first zone contains also one or more of iodine,magnesium, nicotinamide, folic acid and selenium as a said at least oneother active ingredient.
 3. A tablet as claimed in claim 1, wherein saidfirst zone is free from amounts deleterious to the viability of theprobiotic micro-organisms of iron, copper, vitamin B6, vitamin C, zinc,chromium or pantothenic acid or its salts.
 4. (canceled)
 5. (canceled)6. (canceled)
 7. (canceled)
 8. (canceled)
 9. (canceled)
 10. (canceled)11. A tablet as claimed in claim 1, wherein said second zone containsiron, vitamin B6, vitamin C, copper, manganese, pantothenic acid or asalt thereof, zinc or chromium as at least one said other activeingredient.
 12. (canceled)
 13. (canceled)
 14. (canceled)
 15. (canceled)16. (canceled)
 17. (canceled)
 18. (canceled)
 19. A tablet as claimed inclaim 1, wherein said second zone contains any two or more of iron,vitamin B6, vitamin C, pantothenic acid or a salt thereof, zinc, copper,chromium and manganese, each as at least one said other activeingredient.
 20. A tablet as claimed in claim 1, wherein the probioticmicro-organism is mixed with a desiccant carrier material serving toreduce the water activity of the zone containing the probioticmicro-organism, wherein said desiccant carrier material comprises atleast one of carboxymethylcellulose, colloidal silica,polyvinylpyrrolidone, starch, gelatine,hydroxypropylcellulose—low-substituted, microcrystalline cellulose,fumed silicon dioxide, sodium croscarmellose, crospovidone, povidone,magnesium aluminum silicate, methylcellulose, sodium alginate, sodiumstarch glyconate, gelatine, pregelatinized starch, or sorbitol.
 21. Atablet as claimed in claim 1, wherein the second zone contains adesiccant carrier material serving to reduce the water activity of thezone containing the probiotic micro-organism, wherein said desiccantcarrier material comprises at least one of carboxymethylcellulose,colloidal silica, polyvinylpyrrolidone, starch, gelatine,hydroxypropylcellulose—low-substituted, microcrystalline cellulose,fumed silicon dioxide, sodium croscarmellose, crospovidone, povidone,magnesium aluminum silicate, methylcellulose, sodium alginate, sodiumstarch glyconate, gelatine, pregelatinized starch, or sorbitol. 22.(canceled)
 23. A tablet as claimed in claim 1, having a multilayerstructure comprising at least two layers, one of said layersconstituting said first zone and another of said layers constitutingsaid second zone.
 24. A tablet as claimed in claim 23, wherein saidlayer constituting said first zone is free of encapsulated iron, zinc orcopper.
 25. (canceled)
 26. (canceled)
 27. A tablet as claimed in claim1, wherein said layer constituting said first zone contains encapsulatedvitamin B1, vitamin B6, manganese, vitamin A, vitamin D, vitamin E,vitamin B12 or vitamin B2.
 28. (canceled)
 29. A tablet as claimed inclaim 1, wherein said layer constituting said second zone containsencapsulated zinc.
 30. (canceled)
 31. (canceled)
 32. A tablet as claimedin claim 1, having a multitude of granules constituting said first zonesurrounded by a matrix, and wherein said matrix constitutes said secondzone or wherein said matrix also contains a multitude of granulesconstituting said second zone.
 33. A tablet as claimed in claim 1,wherein the water content of the tablet is at least 1% by weight. 34.(canceled)
 35. (canceled)
 36. (canceled)
 37. (canceled)
 38. (canceled)39. A tablet as claimed in claim 1, wherein the water content of thetablet is no more than 7% by weight.
 40. A tablet as claimed in claim 1,wherein the water activity of said first zone is no greater than 0.15.41. (canceled)
 42. (canceled)
 43. (canceled)
 44. (canceled)
 45. A tabletas claimed in claim 1, wherein said first zone is separated from saidsecond zone by a water excluding barrier material, wherein said barriermaterial is a fat or wax based barrier material.
 46. A tablet as claimedin claim 1, wherein the tablet is surrounded by a water excludingbarrier material, wherein said barrier material is a fat or wax basedbarrier material.
 47. (canceled)
 48. (canceled)
 49. (canceled)
 50. Atablet as claimed in claim 1, wherein upon immersion in water a first ofsaid zones disintegrates at a faster rate than a second of said zonessuch that the time for disintegration of said faster disintegrating zoneas a percentage of the total time for disintegration of the faster andthe slower disintegrating zones is no more than 50%.